The objective of this research application is to unveil the molecular and cellular mechanisms by which prostate carcinoma-produced platelet-derived growth factor (PDGF) regulates osteoclastic bone resorption and new bone growth, creating a favorable microenvironment for metastatic deposit. Increasing evidence indicate the significance of PDGF receptor signaling, especially 2-PDGFR, in prostate cancer progression and bone metastasis. PDGF B, however, originally thought to be the sole ligand for 2-PDGFR, has rarely been found in prostate cancer clinical samples. Importantly, our recent immunohistochemical analysis of human prostate carcinoma specimens showed that increased expression of PDGF D, a newly discovered ligand for PDGF receptor-beta (2-PDGFR), is associated with increased Gleason scores and tumor stages. Whereas the classic PDGF ligands A and B are secreted as active dimers, PDGF D contains an N-terminal CUB domain and a C terminal PDGF domain which is secreted as a latent dimer. Extracellular proteolytic cleavage of the CUB domain is required for the PDGF domain to stimulate 2-PDGFR. Recently, we made a novel finding that human prostate carcinoma cells auto-activate latent PDGF D into an active growth factor domain of PDGF D utilizing the serine proteases uPA and matriptase. Consistent with previous reports that matriptase is frequently overexpressed in prostate cancer, especially in metastatic samples, our preliminary data showed increased matriptase expression in human prostate cancer. In a tibiae-injection model, PDGF D significantly enhanced tumor take and growth rate of LNCaP tumors with increased osteolytic and osteoblastic responses. Additionally, our in vitro study unveiled exciting new roles of PDGF D in the regulation of osteoclast differentiation independent of the RANKL/RANK signaling axis as well as osteoblast differentiation involving modulation of homeoprotein expression. Taken together, we hypothesize that prostate carcinoma-produced PDGF D, activated by the transmembrane serine protease matriptase, induces paracrine cell signaling in bone stromal cells, mediating tumor-stromal interactions critical for prostate carcinoma growth in the bone microenvironment. To test this hypothesis, we propose (1) To investigate matriptase-mediated proteolytic processing of PDGF D; (2) To investigate the molecular mechanisms by which PDGF D induces osteoclast and osteoblast differentiation using in vitro models of RAW264.7 and MC3T3-E1 cell lines; and (3) To investigate the roles of PDGF D and its functional regulator matriptase in bone stromal responses and prostate tumor growth in animals. Considering that 2-PDGFR as well as matriptase is highly upregulated in both bone metastases and primary prostate cancer specimens, the completion of the proposed study of interplay between PDGF D, a specific activator of 2-PDGFR, and matriptase will help us in understanding the molecular basis for prostate cancer progression. In addition, the proposed study may provide important information with therapeutic value, especially given that VEGF/PDGF inhibitors are currently in clinical trials for cancer patients. PUBLIC HEALTH RELEVANCE: Mesenchymal growth factors, produced by tumor cells, activate their cognate receptors expressed in surrounding stromal cells, mediating tumor-stromal interactions. These interactions are thought to be critical for tumor progression at both the primary and the metastatic sites. A vast majority of prostate cancer (PC) metastases occur at skeletal sites. The mechanisms underlying the marked propensity of PC to metastasize to bone are beginning to be understood. Although still debatable, it is believed that cell surface and secreted factors produced by the tumor cells induce an osteoclastic response, a critical initial step for PC growth in bones. Bone resorption by osteoclasts results in the release of growth factors and cytokines from the bone matrix, providing a favorable microenvironment for tumor cell proliferation and invasion. Bone formation follows or accompanies bone destruction, and these bone reactions eventually become extensive enough to be visible on radiographs as both sclerotic and lytic reactions. Increasing evidence suggests that platelet-derived growth factor (PDGF) mediates tumor-stromal interactions during human prostate cancer bone metastasis. The goal of this project is to investigate the molecular mechanisms by which prostate carcinoma-produced PDGF initiates bone remodeling, a critical event for tumor cell colonization to the bone. In this application, we will test our hypothesis that the tumor-associated serine proteinase, matriptase, regulates the biological activity of the newly discovered PDGF ligand, PDGF D, and that tumor-derived PDGF D regulates osteoclast differentiation as well as osteoblast proliferation/differentiation critical for tumor cell growth.